Development of endothelial-targeted CD39 as a therapy for ischemic stroke.

BACKGROUND: Ischemic stroke is characterized by a necrotic lesion in the brain surrounded by an area of dying cells termed the penumbra. Salvaging the penumbra either with thrombolysis or mechanical retrieval is the cornerstone of stroke management. At-risk neuronal cells release extracellular adenosine triphosphate, triggering microglial activation and causing a thromboinflammatory response, culminating in endothelial activation and vascular disruption. This is further aggravated by ischemia-reperfusion injury that follows all reperfusion therapies. The ecto-enzyme CD39 regulates extracellular adenosine triphosphate by hydrolyzing it to adenosine, which has antithrombotic and anti-inflammatory properties and reverses ischemia-reperfusion injury. OBJECTIVES: The objective off the study was to determine the efficacy of our therapeutic, anti-VCAM-CD39 in ischaemic stroke. METHODS: We developed anti-VCAM-CD39 that targets the antithrombotic and anti-inflammatory properties of recombinant CD39 to the activated endothelium of the penumbra by binding to vascular cell adhesion molecule (VCAM)-1. Mice were subjected to 30 minutes of middle cerebral artery occlusion and analyzed at 24 hours. Anti-VCAM-CD39 or control agents (saline, nontargeted CD39, or anti-VCAM-inactive CD39) were given at 3 hours after middle cerebral artery occlusion. RESULTS: Anti-VCAM-CD39 treatment reduced neurologic deficit; magnetic resonance imaging confirmed significantly smaller infarcts together with an increase in cerebrovascular perfusion. Anti-VCAM-CD39 also restored blood-brain barrier integrity and reduced microglial activation. Coadministration of anti-VCAM-CD39 with thrombolytics (tissue plasminogen activator [tPA]) further reduced infarct volumes and attenuated blood-brain barrier permeability with no associated increase in intracranial hemorrhage. CONCLUSION: Anti-VCAM-CD39, uniquely targeted to endothelial cells, could be a new stroke therapy even when administered 3 hours postischemia and may further synergize with thrombolytic therapy to improve stroke outcomes.

Advance Care Planning Practice Patterns Before and During a Pandemic.

Introduction: The COVID-19 pandemic resulted in introspection of the United States health care infrastructure, especially with advance care planning (ACP). Methods: This is a retrospective chart review assessing the frequency of ACP discussions and formal documents reflecting ACP wishes in an outpatient palliative medicine (PM) practice. The study site was at University Medical Center New Orleans from pre-COVID-19 surge (November 2019-February 2020) to months during and post-COVID-19 surge (March-April 2020). Results: Results showed an increase in ACP discussions during the post-COVID-19 months. Patients seen during the surge and post-COVID-19 months were more likely to discuss medical power of attorney (odds ratio [OR] = 1.78, p = 0.045) and preferred code status (OR = 2.82, p < 0.001). Conclusion: Our study showed that more ACP discussions were conducted post-COVID-19 versus pre-COVID-19. However, formal documents reflecting these wishes were lacking. These results can help guide ACP use in crisis periods and improve understanding of ACP discussions in an outpatient PM clinic.

Perceptions of Cross-Cultural Challenges and Successful Approaches in Facilitating the Improvement of Equine Welfare.

Projects that aim to improve the welfare of equids worldwide usually involve people from different countries and cultures working together. Given that professionals involved with multi-stakeholder projects often work cross-culturally, this study examined their experiences regarding the challenges involved in, and their reflections on, how to work in a culturally sensitive way. Semi-structured interviews were conducted with 14 participants working in a total of 29 countries and analysed using thematic analysis. Key response themes emerged from the responses to questions covering the areas of perceptions of animal welfare, challenges working cross-culturally and embracing cultural sensitivity. The overriding theme regarding perceptions of animal welfare was that of barriers to animal welfare, under which emerged the subthemes of limited financial and material resources, limited understanding of the tenets of animal welfare, and attachment to traditional medicines and practices. Exploring the key challenges resulted in two themes: challenges regarding the local context and etiquette, and those regarding working with different stakeholders. Considering cultural sensitivity, again, two themes emerged: the importance of trust and respect, and of working with local partners. Previous works have highlighted the importance of shared linguistic knowledge, interpersonal skills and cultural knowledge, and these elements also emerged in this research. As well as providing insights into the challenges of working cross-culturally, the findings of this study have enabled the development of suggestions for how this work could be taken forward in a practical way to be of use to professionals in this sector.

The dermatologist identity crisis: a phenomenological analysis of dermatology trainee professional identity during generalist redeployment.

BACKGROUND: Dermatology faces a workforce crisis against a backdrop of wider medical education shifts towards generalism. A pivot towards generalism may have an impact on dermatology trainee professional identity, which, to our knowledge, has not been investigated and is known to have an impact on competence and conceptualization of ethics among physicians. Disruption because of the COVID-19 pandemic led to dermatology trainee redeployment and therefore presents a unique opportunity to examine dermatology trainee professional identity during times of pressure. AIM: To identify the impact of COVID-19 redeployment on dermatology trainee professional identity, the factors influencing identity and whether such factors affect trainees' perceptions about their future careers. METHODS: Ten trainees were purposively recruited to the study. Data were collected through semistructured interviews and were analysed phenomenologically using the template analysis method. Central and integrative themes were identified. RESULTS: Three central themes were identified: trainee identity and values; redeployment transitions; and future clarity. Three integrative themes were found: tribes; sense of purpose; and uncertainty. Trainees see competence and community as central to the dermatologist identity. Most trainees experienced a reaffirmation of their prior values, psychologically retreating to close-knit communities. However, some underwent disorienting dilemmas, later reflecting on their futures as dermatologists. CONCLUSION: Healthcare crises may have a significant impact on dermatology trainee professional identity and may lead to reaffirmation of prior tribal values or disorienting reflection upon future career trajectories. Improved support among diverse communities, minimization of uncertainty and fostering a sense of purpose among trainees may aid identity enrichment and safeguard the future dermatology consultant workforce.

Under-screened Aboriginal and Torres Strait Islander women's perspectives on cervical screening.

Aboriginal and Torres Strait Islander women have lower participation in Australia's National Cervical Screening Program than other Australian women. Under-screened (including never screened) women's voices are rarely heard in research evidence, despite being a priority group for interventions to increase cervical screening participation. This study aimed to describe under-screened Aboriginal and Torres Strait Islander women's perspectives on cervical screening. Participants were 29 under-screened (women who had either never screened, had not screened in the previous five years or had recently screened in the past three months after more than five years) Aboriginal and Torres Strait Islander women from five communities across three states/territories. Female Aboriginal and Torres Strait Islander researchers Yarned with women about why they did not participate in screening and how to improve screening. Yarning is an Indigenous qualitative research method in which relationships and trust facilitate culturally safe conversation. Transcripts were analysed thematically. The proportion of eligible women who screened within 30 days after the Yarn was calculated. We identified four themes describing how the harms outweighed the benefits of cervical screening for under-screened women. These were: 1) distress, discomfort, and trauma; 2) lack of privacy and control; 3) complicated relationships with health care providers (HCPs); and 4) pressured, insensitive, and/or culturally unsafe communication from HCPs. Under-screened women who had recently screened had maintained privacy and control through self-collection and had experienced trauma-informed and empathetic care from their HCPs. While we cannot unequivocally attribute women's subsequent participation in screening to their involvement in this study, it is notable that one third of eligible under-screened women were screened within 30 days after the Yarn. Enhancing privacy, implementing trauma-informed approaches to care and sensitivity to the clinician-client relationship dynamics could enhance women's sense of comfort in, and control over, the screening procedure. The opportunity to Yarn about cervical screening and self-collection may address these issues and support progress toward cervical cancer elimination in Australia.

CD39 in the development and progression of pulmonary arterial hypertension.

Pulmonary arterial hypertension (PAH) is a devastating progressive disease characterised by pulmonary arterial vasoconstriction and vascular remodelling. Endothelial dysfunction has emerged as a contributing factor in the development of PAH. However, despite progress in the understanding of the pathophysiology of this disease, current therapies fail to impact upon long-term outcomes which remain poor in most patients. Recent observations have suggested the disturbances in the balance between ATP and adenosine may be integral to the vascular remodelling seen in PAH. CD39 is an enzyme important in regulating these nucleos(t)ides which may also provide a novel pathway to target for future therapies. This review summarises the role of adenosine signalling in the development and progression of PAH and highlights the therapeutic potential of CD39 for treatment of PAH.

Aboriginal and Torres Strait Islander women's views of cervical screening by self-collection: a qualitative study.

OBJECTIVE: This study aimed to describe Aboriginal and Torres Strait Islander women's views of self-collection introduced in the renewed National Cervical Screening Program. METHODS: A total of 79 Aboriginal and/or Torres Strait Islander women (50 screened in previous five years, 29 under-screened) from five clinics across three Australian states/territories participated. Topics discussed were perceptions of self-collection, the instruction card and suggestions for implementing self-collection. We employed yarning (a qualitative method), which established relationships and trust between participants and researchers to facilitate culturally safe conversations. Transcripts were analysed thematically. RESULTS: Most women were unaware of self-collection before the yarn but found it to be an acceptable way to participate in cervical screening. Women perceived self-collection would be convenient, provide a sense of control over the screening experience, and maintain privacy and comfort. The instructions were perceived to be simple and easy to follow. Women had concerns about collecting the sample correctly and the accuracy of the sample (compared to clinician-collected samples). CONCLUSIONS: Self-collection is acceptable to Aboriginal and Torres Strait Islander women. IMPLICATIONS FOR PUBLIC HEALTH: Given the inequitable burden of cervical cancer experienced by Aboriginal and Torres Strait Islander women, self-collection is likely to significantly improve participation and ultimately improve cervical cancer outcomes.

Early Endothelial Activation in a Mouse Model of Graft vs Host Disease Following Chemotherapy.

Allogenic hematopoietic stem cell transplant (allo-HSCT) can lead to sinusoidal obstruction syndrome (SOS) and graft-versus-host disease (GvHD) in some individuals. GvHD is characterised by an immune triggered response that arises due to donor T cells recognizing the recipient tissue as "foreign". SOS results in impaired liver function due to microvascular thrombosis and consequent obstruction of liver sinusoids. Endothelial damage occurs following chemotherapy and allo-HSCT and is strongly associated with GvHD onset as well as hepatic SOS. Animal models of GvHD are rarely clinically relevant, and endothelial dysfunction remains uncharacterised. Here we established and characterised a clinically relevant model of GvHD wherein Balb/C mice were subjected to myeloablative chemotherapy followed by transplantation of bone marrow (BM) cells± splenic T-cells from C57Bl6 mice, resulting in a mismatch of major histocompatibility complexes (MHC). Onset of disease indicated by weight loss and apoptosis in the liver and intestine was discovered at day 6 post-transplant in mice receiving BM+T-cells, with established GvHD detectable by histology of the liver within 3 weeks. Together with significant increases in pro-inflammatory cytokine gene expression in the liver and intestine, histopathological signs of GvHD and a significant increase in CD4+ and CD8+ effector and memory T-cells were seen. Endothelial activation including upregulation of vascular cell adhesion molecule (VCAM)- 1 and downregulation of endothelial nitric oxide synthase (eNOS) as well as thrombosis in the liver indicated concomitant hepatic SOS. Our findings confirm that endothelial activation is an early sign of acute GvHD and SOS in a clinically relevant mouse model of GvHD based on myeloablative chemotherapy. Preventing endothelial activation may be a viable therapeutic strategy to prevent GvHD.

Role of Purinergic Signalling in Endothelial Dysfunction and Thrombo-Inflammation in Ischaemic Stroke and Cerebral Small Vessel Disease.

The cerebral endothelium is an active interface between blood and the central nervous system. In addition to being a physical barrier between the blood and the brain, the endothelium also actively regulates metabolic homeostasis, vascular tone and permeability, coagulation, and movement of immune cells. Being part of the blood-brain barrier, endothelial cells of the brain have specialized morphology, physiology, and phenotypes due to their unique microenvironment. Known cardiovascular risk factors facilitate cerebral endothelial dysfunction, leading to impaired vasodilation, an aggravated inflammatory response, as well as increased oxidative stress and vascular proliferation. This culminates in the thrombo-inflammatory response, an underlying cause of ischemic stroke and cerebral small vessel disease (CSVD). These events are further exacerbated when blood flow is returned to the brain after a period of ischemia, a phenomenon termed ischemia-reperfusion injury. Purinergic signaling is an endogenous molecular pathway in which the enzymes CD39 and CD73 catabolize extracellular adenosine triphosphate (eATP) to adenosine. After ischemia and CSVD, eATP is released from dying neurons as a damage molecule, triggering thrombosis and inflammation. In contrast, adenosine is anti-thrombotic, protects against oxidative stress, and suppresses the immune response. Evidently, therapies that promote adenosine generation or boost CD39 activity at the site of endothelial injury have promising benefits in the context of atherothrombotic stroke and can be extended to current CSVD known pathomechanisms. Here, we have reviewed the rationale and benefits of CD39 and CD39 therapies to treat endothelial dysfunction in the brain.

Characterization of a novel model of global forebrain ischaemia-reperfusion injury in mice and comparison with focal ischaemic and haemorrhagic stroke.

Stroke is caused by obstructed blood flow (ischaemia) or unrestricted bleeding in the brain (haemorrhage). Global brain ischaemia occurs after restricted cerebral blood flow e.g. during cardiac arrest. Following ischaemic injury, restoration of blood flow causes ischaemia-reperfusion (I/R) injury which worsens outcome. Secondary injury mechanisms after any stroke are similar, and encompass inflammation, endothelial dysfunction, blood-brain barrier (BBB) damage and apoptosis. We developed a new model of transient global forebrain I/R injury (dual carotid artery ligation; DCAL) and compared the manifestations of this injury with those in a conventional I/R injury model (middle-cerebral artery occlusion; MCAo) and with intracerebral haemorrhage (ICH; collagenase model). MRI revealed that DCAL produced smaller bilateral lesions predominantly localised to the striatum, whereas MCAo produced larger focal corticostriatal lesions. After global forebrain ischaemia mice had worse overall neurological scores, although quantitative locomotor assessment showed MCAo and ICH had significantly worsened mobility. BBB breakdown was highest in the DCAL model while apoptotic activity was highest after ICH. VCAM-1 upregulation was specific to ischaemic models only. Differential transcriptional upregulation of pro-inflammatory chemokines and cytokines and TLRs was seen in the three models. Our findings offer a unique insight into the similarities and differences in how biological processes are regulated after different types of stroke. They also establish a platform for analysis of therapies such as endothelial protective and anti-inflammatory agents that can be applied to all types of stroke.

Strategy for 90% autoverification of clinical chemistry and immunoassay test results using six sigma process improvement.

Six Sigma involves a structured process improvement strategy that places processes on a pathway to continued improvement. The data presented here summarizes a project that took three clinical laboratories from autoverification processes that allowed between about 40% to 60% of tests being auto-verified to more than 90% of tests and samples auto-verified. The project schedule, metrics and targets, a description of the previous system and detailed information on the changes made to achieve greater than 90% auto-verification is presented for this Six Sigma DMAIC (Design, Measure, Analyze, Improve, Control) process improvement project.

Autoverification process improvement by Six Sigma approach: Clinical chemistry & immunoassay.

OBJECTIVE: This study examines effectiveness of a project to enhance an autoverification (AV) system through application of Six Sigma (DMAIC) process improvement strategies. DESIGN AND METHODS: Similar AV systems set up at three sites underwent examination and modification to produce improved systems while monitoring proportions of samples autoverified, the time required for manual review and verification, sample processing time, and examining characteristics of tests not autoverified. This information was used to identify areas for improvement and monitor the impact of changes. RESULTS: Use of reference range based criteria had the greatest impact on the proportion of tests autoverified. To improve AV process, reference range based criteria was replaced with extreme value limits based on a 99.5% test result interval, delta check criteria were broadened, and new specimen consistency rules were implemented. Decision guidance tools were also developed to assist staff using the AV system. The mean proportion of tests and samples autoverified improved from <62% for samples and <80% for tests, to >90% for samples and >95% for tests across all three sites. The new AV system significantly decreased turn-around time and total sample review time (to about a third), however, time spent for manual review of held samples almost tripled. There was no evidence of compromise to the quality of testing process and <1% of samples held for exceeding delta check or extreme limits required corrective action. CONCLUSIONS: The Six Sigma (DMAIC) process improvement methodology was successfully applied to AV systems resulting in an increase in overall test and sample AV by >90%, improved turn-around time, reduced time for manual verification, and with no obvious compromise to quality or error detection.

Women in the Boardroom: Reflecting the Changing Gender Demographics of Dentists.

Even though there are more women in the workplace today than ever before and more women than men are now graduating from college, there are relatively few women who hold leadership positions, be it in government, academics or on corporate boards.

Multiple myeloma presenting in association with gastric phytobezoar.

We present a rare case of a patient with delayed gastric emptying, gastric phytobezoar formation, and osteosclerotic bone lesions as an atypical association with multiple myeloma. Associated gastric features in myeloma, which include diffuse infiltration, gastric plasmacytomas, or delayed gastric emptying, are rare and have a poor prognosis.

Selective inhibition of brain endothelial Rho-kinase-2 provides optimal protection of an in vitro blood-brain barrier from tissue-type plasminogen activator and plasmin.

Rho-kinase (ROCK) inhibition, broadly utilised in cardiovascular disease, may protect the blood-brain barrier (BBB) during thrombolysis from rt-PA-induced damage. While the use of nonselective ROCK inhibitors like fasudil together with rt-PA may be hindered by possible hypotensive side-effects and inadequate capacity to block detrimental rt-PA activity in brain endothelial cells (BECs), selective ROCK-2 inhibition may overcome these limitations. Here, we examined ROCK-2 expression in major brain cells and compared the ability of fasudil and KD025, a selective ROCK-2 inhibitor, to attenuate rt-PA-induced BBB impairment in an in vitro human model. ROCK-2 was highly expressed relative to ROCK-1 in all human and mouse brain cell types and particularly enriched in rodent brain endothelial cells and astrocytes compared to neurons. KD025 was more potent than fasudil in attenuation of rt-PA- and plasminogen-induced BBB permeation under normoxia, but especially under stroke-like conditions. Importantly, only KD025, but not fasudil, was able to block rt-PA-dependent permeability increases, morphology changes and tight junction degradation in isolated BECs. Selective ROCK-2 inhibition further diminished rt-PA-triggered myosin phosphorylation, shape alterations and matrix metalloprotease activation in astrocytes. These findings highlight ROCK-2 as the key isoform driving BBB impairment and brain endothelial damage by rt-PA and the potential of KD025 to optimally protect the BBB during thrombolysis.

Decision dilemmas and opportunities for the new graduate dentist.

Career options for new graduate dentists are changing because of economic and management challenges in traditional practice, the growing trend toward group practice models, enormous educational debt load and lifestyle expectations of the millennial dentist. While new dentists learn to survive and adapt to an evolving profession with many pitfalls, they also have the opportunity to shape the future of our profession.

Pharyngostomy tubes for gastric conduit decompression.

OBJECTIVE: This article illustrates our operative technique for pharyngostomy tube placement and describes our clinical experience with pharyngostomy use for gastric conduit decompression after esophagectomy. METHODS: We retrospectively reviewed patients undergoing pharyngostomy tube placement for gastric conduit decompression after esophagectomy from January 2008 to August 2009. Patients were included if they had a pharyngostomy tube placed at esophagectomy (prophylactic placement) or as a means of decompression after postesophagectomy anastomotic leak (therapeutic placement). We collected operative and clinical data and performed a descriptive statistical analysis. RESULTS: We placed 25 pharyngostomy tubes for gastric conduit decompression after esophagectomy. Eleven were placed prophylactically (44%); the remaining 14 were placed therapeutically (56%) after anastomotic leak. Prophylactic pharyngostomy tubes remained in place a median of 8 days (range 4-17 days), whereas therapeutic pharyngostomy tubes were left in place a median of 15 days (range 7-125 days). There were 4 infectious complications (16%) unrelated to length of pharyngostomy use: 2 cases of cellulitis (resolved with antibiotics, tube remaining in place) and 2 superficial abscesses after tube removal requiring bedside débridement. Seventy-two percent of patients underwent swallow evaluation; 22% of these patients had radiographic evidence of aspiration. CONCLUSIONS: Pharyngostomy tube placement for gastric conduit decompression after esophagectomy is simple, and tubes can stay in place for prolonged periods. Our experience suggests that pharyngostomy tubes are a safe alternative to nasogastric drainage.